The long term objectives of this research proposal are to characterize the signaling pathways downstream of the tyrosine kinase Jak2 which are critical for hematopoietic cell development. That Jak2 plays a critical role in hematopoiesis is demonstrated by the complete absence of several myeloid lineages in mice lacking a functional JAK2 gene. Furthermore, its importance in hematological development is underscored by the recent discovery that leukemic cells of several patients with different hematological malignancies contain an abnormal fusion protein comprised of the transcription factor TEL fused to the catalytic domain of JAK2. Although the signaling cascades activated by Jak2 are not well characterized, recent data suggests that tyrosine autophosphorylation of Jak2 is functionally important. One critical goal of this proposal is to analyze the autophosphorylation sites of JAK2. This will be accomplished, in part, by reconstituting Jak2-/- fetal liver cells with either wild-type or tyrosine-to-phenylalanine point mutants of Jak2, and functionally analyzing the cells by in vitro colony formation or in vivo reconstitution ability. Another important goal will be to identify and characterize molecules which interact directly with phosphorylated Jak2. Such molecules are good candidate Jak2 effector molecules. Thus, this study approaches the problem of studying Jak2 signaling on two fronts: firstly by analyzing Jak2 itself, and secondly, by analyzing the molecules which interact with Jak2.